Promoter and 3'-untranslated-region haplotypes in the vitamin d receptor gene predispose to osteoporotic fracture: the rotterdam study

Polymorphisms of the vitamin D receptor gene (VDR) have been shown to be associated with several complex diseases, including osteoporosis, but the mechanisms are unknown and study results have been inconsistent. We therefore determined sequence variation across the major relevant parts of VDR, including construction of linkage disequilibrium blocks and identification of haplotype alleles. We analyzed 15 haplotype-tagging SNPs in relation to 937 clinical fractures recorded in 6,148 elderly whites over a follow-up period of 7.4 years. Haplotype alleles of the 5' 1a/1e, 1b promoter region and of the 3' untranslated region (UTR) were strongly associated with increased fracture risk. For the 16% of subjects who had risk genotypes at both regions, their risk increased 48% for clinical fractures (P = .0002), independent of age, sex, height, weight, and bone mineral density. The population-attributable risk varied between 1% an

Short-Term, Combined Fasting and Exercise Improves Body Composition in Healthy Males

Fasting enhances the beneficial metabolic outcomes of exercise; however, it is unknown whether body composition is favorably modified on the short term. A baseline-follow-up study was carried out to assess the effect of an established protocol involving short-term combined exercise with fasting on body composition. One hundred seven recreationally exercising males underwent a 10-day intervention across 15 fitness centers in the Netherlands involving a 3-day gradual decrease of food intake, a 3-day period with extremely low caloric intake, and a gradual 4-day increase to initial caloric intake, with daily 30-min submaximal cycling. Using dual-energy X-ray absorptiometry analysis, all subjects substantially lost total body mass (-3.9 ± 1.9 kg; p < .001) and fat mass (-3.3 ± 1.3 kg; p < .001). Average lean mass was lost (-0.6 ± 1.5 kg; p < .001), but lean mass as a percentage of total body mass was not reduced. The authors observed a loss of -3.9 ± 1.9% android fat over total fat mass (p < .001), a loss of -2.2 ± 1.9% gynoid over total fat mass (p < .001), and reduced android/gynoid ratios (-0.05 ± 0.1; p < .001). Analyzing 15 preselected single-nucleotide polymorphisms in 13 metabolism-related genes revealed trending associations for thyroid state-related single-nucleotide polymorphisms rs225014 (deiodinase 2) and rs35767 (insulin-like growth factor1), and rs1053049 (PPARD). In conclusion, a short period of combined fasting and exercise leads to a substantial loss of body and fat mass without a loss of lean mass as a percentage of total mass

Occupational exposure to gases/fumes and mineral dust affect DNA methylation levels of genes regulating expression

Many workers are daily exposed to occupational agents like gases/fumes, mineral dust or biological dust, which could induce adverse health effects. Epigenetic mechanisms, such as DNA methylation, have been suggested to play a role. We therefore aimed to identify differentially methylated regions (DMRs) upon occupational exposures in never-smokers and investigated if these DMRs associated with gene expression levels. To determine the effects of occupational exposures independent of smoking, 903 never-smokers of the LifeLines cohort study were included. We performed three genome-wide methylation analyses (Illumina 450 K), one per occupational exposure being gases/fumes, mineral dust and biological dust, using robust linear regression adjusted for appropriate confounders. DMRs were identified using comb-p in Python. Results were validated in the Rotterdam Study (233 never-smokers) and methylation-expression associations were assessed using Biobank-based Integrative Omics Study data (n = 2802). Of the total 21 significant DMRs, 14 DMRs were associated with gases/fumes and 7 with mineral dust. Three of these DMRs were associated with both exposures (RPLP1 and LINC02169 (2x)) and 11 DMRs were located within transcript start sites of gene expression regulating genes. We replicated two DMRs with gases/fumes (VTRNA2-1 and GNAS) and one with mineral dust (CCDC144NL). In addition, nine gases/fumes DMRs and six mineral dust DMRs significantly associated with gene expression levels. Our data suggest that occupational exposures may induce differential methylation of gene expression regulating genes and thereby may induce adverse health effects. Given the millions of workers that are exposed daily to occupational exposures, further studies on this epigenetic mechanism and health outcomes are warranted

Controlling bias and inflation in epigenome- and transcriptome-wide association studies using the empirical null distribution

We show that epigenome- and transcriptome-wide association studies (EWAS and TWAS) are prone to significant inflation and bias of test statistics, an unrecognized phenomenon introducing spurious findings if left unaddressed. Neither GWAS-based methodology nor state-of-the-art confounder adjustment methods completely remove bias and inflation. We propose a Bayesian method to control bias and inflation in EWAS and TWAS based on estimation of the empirical null distribution. Using simulations and real data, we demonstrate that our method maximizes power while properly controlling the false positive rate. We illustrate the utility of our method in large-scale EWAS and TWAS meta-analyses of age and smoking

Mendelian randomization integrating GWAS and eQTL data reveals genetic determinants of complex and clinical traits

Genome-wide association studies (GWAS) have identified thousands of variants associated with complex traits, but their biological interpretation often remains unclear. Most of these variants overlap with expression QTLs, indicating their potential involvement in regulation of gene expression. Here, we propose a transcriptome-wide summary statistics-based Mendelian Randomization approach (TWMR) that uses multiple SNPs as instruments and multiple gene expression traits as exposures, simultaneously. Applied to 43 human phenotypes, it uncovers 3,913 putatively causal gene-trait associations, 36% of which have no genome-wide significant SNP nearby in previous GWAS. Using independent association summary statistics, we find that the majority of these loci were missed by GWAS due to power issues. Noteworthy among these links is educational attainment-associated BSCL2, known to carry mutations leading to a Mendelian form of encephalopathy. We also find pleiotropic causal effects suggestive of mechanistic connections. TWMR better accounts for pleiotropy and has the potential to identify biological mechanisms underlying complex traits

Refining Attention-Deficit/Hyperactivity Disorder and Autism Spectrum Disorder Genetic Loci by Integrating Summary Data From Genome-wide Association, Gene Expression, and DNA Methylation Studies

Background: Recent genome-wide association studies (GWASs) identified the first genetic loci associated with attention-deficit/hyperactivity disorder (ADHD) and autism spectrum disorder (ASD). The next step is to use these results to increase our understanding of the biological mechanisms involved. Most of the identified variants likely influence gene regulation. The aim of the current study is to shed light on the mechanisms underlying the genetic signals and prioritize genes by integrating GWAS results with gene expression and DNA methylation (DNAm) levels. Methods: We applied summary-data–based Mendelian randomization to integrate ADHD and ASD GWAS data with fetal brain expression and methylation quantitative trait loci, given the early onset of these disorders. We also analyzed expression and methylation quantitative trait loci datasets of adult brain and blood, as these provide increased statistical power. We subsequently used summary-data–based Mendelian randomization to investigate if the same variant influences both DNAm and gene expression levels. Results: We identified multiple gene expression and DNAm levels in fetal brain at chromosomes 1 and 17 that were associated with ADHD and ASD, respectively, through pleiotropy at shared genetic variants. The analyses in brain and blood showed additional associated gene expression and DNAm levels at the same and additional loci, likely because of increased statistical power. Several of the associated genes have not been identified in ADHD and ASD GWASs before. Conclusions: Our findings identified the genetic variants associated with ADHD and ASD that likely act through gene regulation. This facilitates prioritization of candidate genes for functional follow-up studies

DNA methylation signatures of aggression and closely related constructs : A meta-analysis of epigenome-wide studies across the lifespan

DNA methylation profiles of aggressive behavior may capture lifetime cumulative effects of genetic, stochastic, and environmental influences associated with aggression. Here, we report the first large meta-analysis of epigenome-wide association studies (EWAS) of aggressive behavior (N = 15,324 participants). In peripheral blood samples of 14,434 participants from 18 cohorts with mean ages ranging from 7 to 68 years, 13 methylation sites were significantly associated with aggression (alpha = 1.2 x 10(-7); Bonferroni correction). In cord blood samples of 2425 children from five cohorts with aggression assessed at mean ages ranging from 4 to 7 years, 83% of these sites showed the same direction of association with childhood aggression (r = 0.74, p = 0.006) but no epigenome-wide significant sites were found. Top-sites (48 at a false discovery rate of 5% in the peripheral blood meta-analysis or in a combined meta-analysis of peripheral blood and cord blood) have been associated with chemical exposures, smoking, cognition, metabolic traits, and genetic variation (mQTLs). Three genes whose expression levels were associated with top-sites were previously linked to schizophrenia and general risk tolerance. At six CpGs, DNA methylation variation in blood mirrors variation in the brain. On average 44% (range = 3-82%) of the aggression-methylation association was explained by current and former smoking and BMI. These findings point at loci that are sensitive to chemical exposures with potential implications for neuronal functions. We hope these results to be a starting point for studies leading to applications as peripheral biomarkers and to reveal causal relationships with aggression and related traits.Peer reviewe

Autosomal genetic variation is associated with DNA methylation in regions variably escaping X-chromosome inactivation

X-chromosome inactivation (XCI), i.e., the inactivation of one of the female X chromosomes, restores equal expression of X-chromosomal genes between females and males. However, ~10% of genes show variable degrees of escape from XCI between females, although little is known about the causes of variable XCI. Using a discovery data-set of 1867 females and 1398 males and a replication sample of 3351 females, we show that genetic variation at three autosomal loci is associated with female-specific changes in X-chromosome methylation. Through cis-eQTL expression analysis, we map these loci to the genes SMCHD1/METTL4, TRIM6/HBG2, and ZSCAN9. Low-expression alleles of the loci are predominantly associated with mild hypomethylation of CpG islands near genes known to variably escape XCI, implicating the autosomal genes in variable XCI. Together, these results suggest a genetic basis for variable escape from XCI and highlight the potential of a population genomics approach to identify genes involved in XCI

Skewed X-inactivation is common in the general female population

X-inactivation is a well-established dosage compensation mechanism ensuring that X-chromosomal genes are expressed at comparable levels in males and females. Skewed X-inactivation is often explained by negative selection of one of the alleles. We demonstrate that imbalanced expression of the paternal and maternal X-chromosomes is common in the general population and that the random nature of the X-inactivation mechanism can be sufficient to explain the imbalance. To this end, we analyzed blood-derived RNA and whole-genome sequencing data from 79 female children and their parents from the Genome of the Netherlands project. We calculated the median ratio of the paternal over total counts at all X-chromosomal heterozygous single-nucleotide variants with coverage ≥10. We identified two individuals where the same X-chromosome was inactivated in all cells. Imbalanced expression of the two X-chromosomes (ratios ≤0.35 or ≥0.65) was observed in nearly 50% of the population. The empirically observed skewing is explained by a theoretical model where X-inactivation takes place in an embryonic stage in which eight cells give rise to the hematopoietic compartment. Genes escaping X-inactivation are expressed from both alleles and therefore demonstrate less skewing than inactivated genes. Using this characteristic, we identified three novel escapee genes (SSR4, REPS2, and SEPT6), but did not find support for many previously reported escapee genes in blood. Our collective data suggest that skewed X-inactivation is common in the general population. This may contribute to manifestation of symptoms in carriers of recessive X-linked disorders. We recommend that X-inactivation results should not be used lightly in the interpretation of X-linked variants